Fig. 2: 3D map of the microvascular environment in 4-weeks-old murine calvaria.
From: Quantitative 3D imaging of the cranial microvascular environment at single-cell resolution
A, B MIP displaying calvarial vessels (Ai, Bi) and Osterix+ (Ai, Ai’) or Gli1+ skeletal progenitors (Bi, Bi’) in the parietal and posterior frontal bones of the calvarium. Zoomed-in images demonstrate the distribution of vessel phenotypes (Aii–vi, Bii–vi) and skeletal progenitors (Aii–vi, Aii’–vi’, Bii–vi, Bii’–vi’) in different regions. C, D 50-μm-thick sagittal sections showing the locations of vessels Ci–ii, Di–ii) and skeletal progenitors (Ci–ii, Ci’–ii’, Di–ii, Di’–ii’) along the thickness of the calvarium. CD31hiEmcn− and CD31hiEmcnhi vessels are prevalent in the periosteum and dura mater (yellow arrows), while CD31hiEmcnhi and CD31loEmcnhi vessels make up the majority of vessels in the bone marrow cavities (white arrows). Osterix+ progenitors (A, C) are distributed throughout the periosteum and dura mater (yellow arrows), transcortical canals (white arrowheads), and osteogenic fronts nearby the sutures (yellow arrowheads), while Gli1+ progenitors (B, D) are mainly restricted to the transcortical canals and sutures. Results were replicated in 3 calvaria for each staining combination (CD31/Emcn/Osterix, CD31/Emcn/Gli1). E Plots showing the nearest distance of Osterix+ (top) and Gli1+ (bottom) progenitors to each vessel phenotype (n = 3). Both cell types exhibit a preferential spatial relationship to CD31hiEmcnhi vessels compared to the other phenotypes. CD31loEmcnlo vessels were not quantified due to their low fluorescence signal-to-noise ratio. Data are mean ± SD. Statistics were performed using a two-way ANOVA and Bonferroni post-hoc test. ***p < 0.001, **p < 0.01, *p < 0.05 between CD31hiEmcnhi and CD31hiEmcn− vessels; +++p < 0.001 between CD31loEmcnhi and CD31hiEmcn− vessels; Scale bars: 1000 μm (Ai, Bi), 100 μm (Aii–vi, Bii–vi), 700 μm (Ci), 300 μm (Cii), 500 μm (Di), 200 μm (Dii). Colors: Red: Endomucin (Emcn), Gray: Osterix (A, C) or Gli1 (B, D), Green: CD31.
